DNA analyzer

ABSTRACT

Aspects of the disclosure provide a microfluidic chip to facilitate DNA analysis. The microfluidic chip includes a first domain configured for polymerase chain reaction (PCR) amplification of DNA fragments, a dilution domain coupled to the first domain to dilute a PCR mixture received from the first domain, and a second domain that is coupled to the dilution domain so as to receive the amplified DNA fragments. The second domain includes a separation channel that is configured to perform electrophoretic separation of the amplified DNA fragments. In addition, the disclosure provides a DNA analyzer to act on the microfluidic chip to perform an integrated single chip DNA analysis.

INCORPORATION BY REFERENCE

This application claims the benefit of U.S. Provisional Applications No.61/213,405, “Fast Sample to Answer DNA Analyzer (AnalyticalMicrodevice)” filed on Jun. 4, 2009, No. 61/213,406, “Optical Approachfor Microfluidic DNA Electrophoresis Detection” filed on Jun. 4, 2009,and No. 61/213,404, “Multiple Sample, Integrated Microfluidic Chip forDNA Analysis” filed on Jun. 4, 2009, which are incorporated herein byreference in their entirety.

BACKGROUND

DNA is recognized as the “ultimate biometric” for human identification.DNA analysis can provide evidence for solving forensic and medicalcases, such as in areas of criminal justice, identifications of humanremains, paternity testing, pathogen detection, disease detection, andthe like.

SUMMARY

Aspects of the disclosure provide a microfluidic chip to facilitate DNAanalysis. The microfluidic chip can be used in a DNA analyzer for anintegrated single chip DNA analysis. The microfluidic chip can include afirst domain configured for polymerase chain reaction (PCR)amplification of DNA fragments, a dilution domain configured to dilute aPCR mixture received from the first domain, and a second domain that iscoupled to the dilution domain so as to receive the amplified DNAfragments. The second domain includes a separation channel that isconfigured to perform electrophoretic separation of the amplified DNAfragments. Additionally, the microfluidic chip can include otherdomains, such as a DNA purification domain, a post-PCR clean-up/dilutiondomain, and the like.

In an example, the first domain can include a reservoir configured toreceive a template DNA and reagents, and generate target DNA fragmentsbased on the template DNA and the reagents. The microfluidic chip caninclude a plurality of inlets to input the template DNA and the reagentsinto the microfluidic chip.

In addition to the separation channel, the second domain can include aninjection channel configured to inject amplified DNA fragments into theseparation channel by electro-kinetic or pressure injection.

The microfluidic chip can also include a plurality of electrodereservoirs for applying electric fields in the separation channel forthe electrophoretic separation of the amplified DNA fragments, and forapplying electric fields in the injection channel for theelectro-kinetic injection. Additionally, the microfluidic chip caninclude a waste outlet to collect waste liquid generated on themicrofluidic chip.

Further, the dilution domain is configured to dilute the PCR mixturereceived from the first domain according to a ratio from 1:5 to 1:20(one part of the PCR mixture to 5-20 parts of a dilutant).

Aspects of the disclosure can also provide a cartridge. The cartridgecan include a sample acceptor configured to extract a template DNA, andthe microfluidic chip. The cartridge can be installed in a DNA analyzerat a time of DNA analysis, and can be thrown away after the DNAanalysis. In addition, the cartridge can include a reagent carrierconfigured to carry reagents required for the PCR amplification, and theelectrophoretic separation.

According to the disclosure, the sample acceptor can extract thetemplate DNA by any suitable solid phase extraction or liquid phaseextraction, such as silica solid phase extraction, liquid phaseenzymatic DNA isolation, and the like. In an example, the sampleacceptor includes a well having a liquid phase mixture to extract thetemplate DNA by liquid phase enzymatic DNA isolation. The liquid phasemixture can be sealed in the well before the extraction.

Aspects of the disclosure can provide a DNA analyzer to perform DNAanalysis using the microfluidic chip. The DNA analyzer can include aninterface for coupling the microfluidic chip to the DNA analyzer, apressure module configured to flow liquid in the microfluidic chip, athermal module configured to induce thermal cycles at the first domainof the microfluidic chip for the PCR amplification, a power moduleconfigured to generate voltages to be applied to the second domain ofthe microfluidic chip for the electrophoretic separation, a detectionmodule configured to excite fluorescent labels attached to DNA fragmentsto emit fluorescence and detect the emitted fluorescence, and acontroller module. The controller module can control the pressuremodule, the thermal module, the power module, and the detection moduleaccording to a control procedure to act on the microfluidic chip for asingle-chip DNA analysis.

In an embodiment, the pressure module includes a plurality of pumpsand/or vacuums configured to inject a template DNA and reagents in themicrofluidic chip. The controller module can respectively control theplurality of pumps and/or vacuums. In another embodiment, themicrofluidic chip has a membrane valve. The pressure module can includea vacuum pump configured to control the membrane valve to enable fluidmovement.

The thermal module includes components to induce thermal cycles for thePCR amplification. In an embodiment, the thermal module includes aheating unit configured to direct heat to the first domain, a coolingunit configured to disperse heat from the first domain, and a sensingunit configured to measure solution temperature in the first domain. Inan example, the heating unit includes an infrared light source to directheat to the first domain, the cooling unit includes a cooling fan, andthe sensing unit includes an infrared pyrometer to measure thetemperature. Thus, the infrared light source, the cooling fan and theinfrared pyrometer can induce the thermal cycles in a reservoir at thefirst domain for the PCR amplification without contacting the reservoir.In another example, the first domain includes a thermal-couplerreservoir coupled with a PCR reservoir for the PCR amplification. Thesensing unit can use any suitable techniques, such as contactingtechniques, non-contacting techniques, and the like, to measure solutiontemperature in the thermal-coupler reservoir. Then, the infrared lightsource, the cooling fan can induce the thermal cycles in the PCRreservoir for the PCR amplification based on temperature measured fromthe thermal-coupler reservoir.

In an embodiment, the detection module includes a laser module, a set ofoptics and a detection module. The laser module is configured to emit alaser beam. The detection module is configured to detect fluorescence.The set of optics is configured to direct the laser beam to theseparation channel to excite the fluorescent labels to emitfluorescence, and direct the emitted fluorescence to the detectionmodule for detection.

Additionally, the DNA analyzer can include a mechanism to identify asample. In an example, the DNA analyzer includes a barcode readerconfigured to read a barcode to identify a sample. In another example,the DNA analyzer includes a radio frequency identification (RFID) readerconfigured to read a RFID tag to identify a sample.

Aspects of the disclosure can provide a method for DNA analysis. Themethod includes inducing thermal cycles in a first domain of amicrofluidic chip for PCR amplification of DNA fragments, inducingliquid flow to move the amplified DNA fragments from the first domain toa second domain of the microfluidic chip having a separation channel forelectrophoretic separation, inducing an electric field over theseparation channel to separate the DNA fragments by size, and detectingthe separated DNA fragments.

Additionally, the method can include injecting reagents into a reservoirin the first domain for the PCR amplification, and injecting copies oftemplate DNA into the reservoir for the PCR amplification.

Further, the method can include diluting the amplified DNA fragments ina dilution solution, and electro-kinetically injecting the DNA fragmentsinto the separation channel.

According to an embodiment of the disclosure, the amplified DNAfragments are tagged with fluorescent labels or intercalated with dyes.Thus, to detect the separated DNA fragments, the method can includeemitting a laser beam, directing the laser beam to the separationchannel to excite the fluorescent labels to emit fluorescence,collecting the emitted fluorescence, and returning the collectedfluorescence for detection.

In an embodiment, to assist size measurement, the method can includeadding an internal lane standard (ILS) into the amplified DNA fragments.

BRIEF DESCRIPTION OF THE DRAWINGS

Various exemplary embodiments of this disclosure will be described indetail with reference to the following figures, wherein like numeralsreference like elements, and wherein:

FIG. 1 shows a block diagram of an exemplary DNA analyzer according toan embodiment of the disclosure;

FIGS. 2A-2C show a swab example and elevation views of a samplecartridge example according to an embodiment of the disclosure;

FIG. 3 shows a schematic diagram of a microfluidic chip exampleaccording to an embodiment of the disclosure;

FIG. 4 shows an exemplary DNA analyzer according to an embodiment of thedisclosure;

FIG. 5 shows a flow chart outlining an exemplary process for using a DNAanalyzer to perform DNA analysis according to an embodiment of thedisclosure;

FIG. 6 shows a flow chart outlining an exemplary process for a DNAanalyzer to perform DNA analysis according to an embodiment of thedisclosure;

FIG. 7 shows a plot of thermal cycles for polymerase chain reaction(PCR) amplification according to an exemplary DNA analyzerimplementation; and

FIG. 8 shows a plot of fluorescent light detection according to anexemplary DNA analyzer implementation.

DETAILED DESCRIPTION OF EMBODIMENTS

FIG. 1 shows a block diagram of an exemplary DNA analyzer 100 accordingto an embodiment of the disclosure. The DNA analyzer 100 includes amicrofluidic chip module 110, a thermal module 120, a pressure module130, a high voltage module 140, a detection module 150, a power module160, a computing module 170, and a controller module 180. Additionally,the DNA analyzer 100 can include a magnetic module 190. These elementscan be coupled together as shown in FIG. 1.

The DNA analyzer 100 is capable of processing sample-to-answer DNAanalysis on an integrated single-chip. Thus, using the DNA analyzer 100to perform DNA analysis does not need substantial experience andknowledge of DNA processes. In an example, the appropriate procedures touse the DNA analyzer 100 to perform DNA analysis can be learned in anhour. Additionally, the integrated single-chip DNA analysis requires areduced volume of reagents, for example, in the order of a micro-liter.Further, the reduced volume of reagents can reduce thermal inputs forinducing thermal cycles in the DNA analysis, and thus reduce the timefor DNA analysis.

The microfluidic chip module 110 includes a microfluidic chip 111. Themicrofluidic chip 111 can be suitably coupled with other elements of theDNA analyzer 100 to perform integrated single-chip DNA analysis. In anexample, the microfluidic chip module 110 is implemented as a disposablecartridge, and a cartridge interface that can couple the disposablecartridge with other components of the DNA analyzer 100 that are notincluded as part of the disposable cartridge. The disposable cartridgeincludes the microfluidic chip 111 and a micro-to-macro interface. Themicro-to-macro interface couples the microfluidic chip 111 to macrostructures on the disposable cartridge. The disposable cartridge can beseparately stored, and can be installed in the DNA analyzer 100 at atime of DNA analysis. After the DNA analysis, the disposable cartridgecan be suitably thrown away.

The microfluidic chip 111 includes various domains that can be suitablyconfigured to enable the integrated single-chip DNA analysis. In anembodiment, DNA analysis generally includes a step of PCR amplification,and a step of electrophoretic separation. The microfluidic chip 111 caninclude a first domain 111 a for the PCR amplification and a seconddomain 111 b for the electrophoretic separation. In addition, themicrofluidic chip 111 can include other domains that are suitablyintegrated with the first domain 111 a and the second domain 111 b. Inan example, the microfluidic chip 111 includes a purification domainfluidically coupled with the first domain 111 a. The purification domaincan be used to extract and purify a template DNA. It is noted that anysuitable techniques, such as solid-phase extraction, liquid-phaseextraction, and the like, can be used to purify the template DNA in thepurification domain.

In another example, the microfluidic chip 111 includes a post-PCRclean-up/dilution domain that is fluidically coupled with the firstdomain 111 a and the second domain 111 b. The post-PCR clean-up/dilutiondomain can be used for any suitable process after the PCR amplificationand before the electrophoretic separation.

The first domain 111 a includes a reservoir configured for PCRamplification. In an embodiment, the first domain 111 a includesmultiple separated reservoirs to enable simultaneous PCR amplificationfor multiple DNA samples. The temperature at the first domain 111 a canbe controlled by the thermal module 120 to enable the PCR amplification.According to an embodiment of the disclosure, the PCR amplification onthe microfluidic chip 111 requires only a small volume of reagents, andthe PCR amplification can achieve rapid thermal cycling. In an example,the volume of reagents used for the PCR amplification can be in theorder of sub-micro-liter, and the time required for the PCRamplification can be under 20 minutes.

The second domain 111 b can include a plurality of micro channels. Theplurality of micro channels can be configured for electrophoreticseparation. More specifically, each micro channel can be filled with,for example, polymer sieving matrix. Further, an electric field can beinduced in the micro channel. Thus, when DNA fragments are injected inthe micro channel, the DNA fragments can migrate by force of theelectric field at different speeds based on the sizes of the DNAfragments.

Additionally, the second domain 111 b can be configured to facilitateDNA fragments detection in the DNA analysis. In an example, DNAfragments are tagged with fluorescent labels during PCR, before beinginjected in the micro channels. The fluorescent labels can emitfluorescence of pre-known wavelength when excited by a laser beam. Thesecond domain 111 b includes a detection window configured fordetection. The laser beam can be directed to pass through the detectionwindow to excite the fluorescent labels in the micro channels. Theemitted fluorescence can pass through the detection window to becollected and detected.

The microfluidic chip 111 can include additional structures tofacilitate the integrated single-chip DNA analysis. For example, themicrofluidic chip 111 can include microfluidic channels that can directDNA fragments from the first domain 111 a to the second domain 111 b.Through the microfluidic channels, the DNA fragments flow in a solutionfrom the first domain 111 a to the second domain 111 b. In addition, themicrofluidic chip 111 can include inlets for receiving reagents and thetemplate DNA. The microfluidic chip 111 can also include additionalreservoirs for additional processing steps, such as dilution, cleanup,and the like.

The microfluidic chip 111 can be constructed from any suitable material.In an example, the microfluidic chip 111 is constructed from glass. Inanother example, the microfluidic chip 111 is constructed from plasticor polymeric material.

In addition to the microfluidic chip 111, the disposable cartridge caninclude a sample acceptor and a reagent carrier. In an example, thesample acceptor accepts a swab used for taking DNA sample, such as fromsaliva, bloodstains, cigarettes, and the like. Further, the sampleacceptor extracts a template DNA from the swab. The sample acceptor canuse any suitable mechanism, such as solid-phase extraction, liquid-phaseextraction, and the like to obtain and/or purify the template DNA fromthe swab. In an embodiment, the sample acceptor uses a solid-phase DNAextraction method, such as silica beads based DNA extraction.

In another embodiment, the sample acceptor uses a liquid-phase DNAextraction method. The liquid-phase DNA extraction method can simplifythe purification and extraction process, and reduce a total cost of theDNA analyzer 100. In an example, the sample acceptor uses an enzymaticDNA-isolation method to extract and purify the template DNA. Theenzymatic DNA-isolation method can achieve liquid phase purificationwithout a need of centrifugation. In addition, the sample acceptor canbe suitably designed to maintain sample integrity.

More specifically, the sample acceptor can include a plurality ofseparated wells for taking swabs, for example. Thus, the DNA analysiscan simultaneously process multiple DNA samples. Each well includes aliquid phase mixture that is sealed by a membrane at a bottom portion ofthe well. The liquid phase mixture can conduct enzymatic digestion ofall proteins and other cellular interferences, with the exception ofDNA. In an embodiment, the liquid phase mixture can include thermostableproteinases from thermophilic Bacillus species. For example, a liquidphase mixture including the thermostable proteinases from thermophilicBacillus species is disclosed in U.S. Patent Application Publication No.2004/0197788, which is incorporated herein by reference in its entirety.Thus, the liquid phase mixture can perform DNA extraction andpurification when a swab is immersed in the liquid phase mixture. Theliquid phase method can achieve comparable DNA quality to othermethodologies in both DNA concentration and purity. In an example, afinal DNA concentration by the liquid phase method is in a range of0.5-2 ng/μL.

Further, using the liquid phase extraction method instead of the silicasolid phase method can reduce the overall hydraulic pressure requirementto induce solution flow through the microfluidic chip 111. In anembodiment, the liquid phase extraction can enable a valveless designfor the microfluidic chip 111. Thus, the liquid phase extraction cansimplify the DNA analyzer 100 and simplify the manufacturing and testingsteps in association with the solid-phase extraction.

Before taking DNA sample, a swab can be sealed in a hard case to avoidcontamination. The swab can be attached to a seal cap that can seal thehard case. The swab can be identified by various mechanisms. In anexample, a barcode label is attached to the hard case to identify theswab. In another example, the seal cap has a radio frequencyidentification (RFID) tag implanted. The RFID tag can identify the swabattached to the seal cap throughout the process. After the swab is usedto take DNA sample, the swab can be placed in one of the plurality ofseparated wells, and can be sealed in the well, for example, by the sealcap attached to the sampled swab. In an embodiment, the seal cap is astepped seal cap that can seal the well in a first step, and a secondstep. When the seal cap seals the well in the first step, the swab doesnot puncture the membrane. When the seal cap seals the well in thesecond step, the swab punctures the membrane and is immersed in theliquid phase mixture. The liquid phase mixture can then extract templateDNA from the swab.

The reagent carrier can house a plurality of reagents for DNA analysis,such as reagents for polymerase chain reaction (PCR) amplification,solutions for electrophoretic separation, and the like. In an STR typingexample, the reagent carrier houses reagents for multiplexed STRamplification. The reagents can perform multiplexed STR amplificationand can use multiple fluorescent dyes to label STR alleles. The reagentscan be commercially available reagent kits or can be tailored to themicro-scale chip environment to further facilitate the integratedsingle-chip DNA analysis.

In addition, the reagent carrier houses solutions that are suitable forelectrophoretic separation in the micro-scale chip environment. Forexample, the reagent carrier houses a coating solution, such aspoly-N-hydroxyethylacrylamide, and the like. The coating solution can beused to coat micro channel walls prior to the separation to reduceelectro osmotic flow and enable single base pair resolution of amplifiedDNA fragments. In another example, the reagent carrier houses a dilutionsolution, such as water and/or Formamide, and the like. The dilutionsolution can be used to reduce the ionic strength of the sample in orderto promote better electro-kinetic injection. In another example, thereagent carrier houses an internal lane standard (ILS). The ILS can beused for accurate size measurements. The reagent carrier also houses apolymer solution for electrophoretic separation in the micro-scale chipenvironment. The polymer solution is used as gels to provide a physicalseparation of DNA fragments according to chain length. In an embodiment,the polymer solution can include a sieving or non-sieving matrix. Forexample, polymer solutions including a sieving or non-sieving matrix aredisclosed in U.S. Pat. No. 7,531,073, No. 7,399,396, No. 7,371,533, No.7,026,414, No. 6,811,977 and No. 6,455,682, which are incorporatedherein by reference in their entirety. In an example, a polymer sievingmatrix can be used to yield a single-base resolution in a totalseparation length of 8 cm and in less than 400 seconds.

The thermal module 120 receives control signals from the controllermodule 180, and induces suitable temperatures for DNA analysis, such asa temperature for DNA extraction, thermal cycles for the PCRamplification, a temperature for electrophoretic separation, and thelike. In an example, the thermal module 120 includes a resistance heaterto control a temperature in the wells of the sample acceptor for the DNAextraction and purification. In another example, the thermal module 120includes another resistance heater to control a temperature at thesecond domain 111 b.

In another example, the thermal module 120 includes a heating unit, acooling unit and a sensing unit to induce the thermal cycles for the PCRamplification at the first domain 111 a. The heating unit can directheat to the first domain 111 a, the cooling unit can disperse heat fromthe first domain 111 a, and the sensing unit can measure a temperatureat the first domain 111 a. The controller module 180 can control theheating unit and the cooling unit based on the temperature measured bythe sensing unit.

In an embodiment, the thermal module 120 performs non-contact thermalcontrols. For example, the thermal module 120 includes an infrared lightsource as the heating unit, a cooling fan as the cooling unit, and aninfrared pyrometer as the temperature sensing unit. The infrared lightsource, such as a halogen light bulb, can excite, for example, the 1.3μm vibrational band of liquid. Thus, the infrared light source can heata small volume of solution within a reservoir in the first domain 111 aindependent of the reservoir to achieve rapid heating and cooling. Theinfrared pyrometer measures blackbody radiation from an outside of thereservoir. In an example, the reservoir is designed to have a thinnerside for the infrared pyrometer measurements. The infrared pyrometermeasurements at the thinner side can more accurately reflect thetemperature of solution within the reservoir. Thus, the DNA analyzer 100can achieve a precise temperature control along with rapid thermalcycles. In an example, the DNA analyzer 100 can achieve a temperaturefluctuation of less than ±0.1° C., and a time of the thermal cycles forthe PCR amplification can be less than 20 minutes.

The pressure module 130 receives control signals from the controllermodule 180, and applies suitable pressures to the microfluidic chipmodule 110 to enable fluid movement. In an embodiment, the pressuremodule 130 receives a sensing signal that is indicative of a pressureapplied to the microfluidic chip module 110, and suitably adjusts itsoperation to maintain the suitable pressure to the microfluidic chipmodule 110.

The pressure module 130 can include a plurality of pumps. The pluralityof pumps control the injection of the various reagents and the templateDNA solutions into the microfluidic chip 111. According to an embodimentof the disclosure, the plurality of pumps can be individually controlledto achieve any possible timing sequence.

The pressure module 130 may include other pressure components to suitthe integrated single-chip integrated DNA analysis. In an embodiment,the microfluidic chip 111 has membrane valves. The pressure module 130can include a hydrodynamic pressure/vacuum system to suitably controlthe closing and opening of the membrane valves to enable fluid movementthrough the microfluidic chip 111.

In another embodiment, the microfluidic chip 111 is valveless. Forexample, the DNA analyzer 100 uses a liquid phase DNA extraction insteadof a silica solid phase DNA extraction. The liquid phase DNA extractioncan be integrated with following DNA processes on a valvelessmicrofluidic chip. Thus, the hydrodynamic pressure/vacuum system is notneeded. The pressure module 130 can be simplified to reduce thefootprint, the weight, the cost, and the complexity of the DNA analyzer100.

The power module 160 receives a main power, and generates variousoperation powers for various components of the DNA analyzer 100. In anexample, the DNA analyzer 100 is implemented using a modular design.Each module of the DNA analyzer 100 needs an operation power supply,which can be different from other modules. The power module 160 receivesan AC power input, such as 100-240 V, 50-60 Hz, single phase AC powerfrom a power outlet. Then, the power module 160 generates 5 V, 12 V, 24V, and the like, to provide operation powers for the various componentsof the DNA analyzer 100.

In addition, the power module 160 generates high voltages, such as 1000V, 2000 V, and the like, for suitable DNA processes on the microfluidicchip 111, such as electro-kinetic injection, electrophoretic separation,and the like.

Further, the power module 160 can implement various protectiontechniques, such as power outrage protection, graceful shut-down, andthe like, to protect the various components and data against powerfailure. It is noted that the power module 160 may include a back-uppower, such as a battery module, to support, for example, gracefulshut-down.

The high voltage module 140 can receive the high voltages from the powermodule 160 and suitably apply the high voltages on the microfluidic chip111. For example, the high voltage module 140 includes interfaces thatapply the high voltages to suitable electrodes on the microfluidic chip111 to induce electro-kinetic injection and/or electrophoreticseparation.

The detection module 150 includes components configured to suit theintegrated single-chip DNA analysis. In an embodiment, the detectionmodule 150 is configured for multicolor fluorescence detection. Thedetection module 150 includes a laser source unit, a set of optics and adetector unit.

The laser source unit emits a laser beam. In an example, the lasersource unit includes an argon-ion laser unit. In another example, thelaser source unit includes a solid state laser, such as a coherentsapphire optically pumped semiconductor laser unit. The solid statelaser has the advantages of reduced size, weight and power consumption.

The set of optics can direct the laser beam to pass through thedetection window at the second domain 111 b of the microfluidic chip111. The laser beam can excite fluorescent labels attached to DNAfragments to emit fluorescence. Further, the set of optics can collectand direct the emitted fluorescence to the detector unit for detection.In an STR typing example, STR alleles are separated in the second domain111 b according to sizes. STR alleles of different sizes pass thedetection window at different times. In addition, STR alleles ofoverlapping sizes can be tagged with fluorescent labels of differentcolors. The detector unit can be configured to detect an STR allelehaving a fluorescent label based on a time of fluorescence emitted bythe fluorescent label and a color of the emitted fluorescence.

In another example, internal lane standard (ILS) is added to migrate inthe micro channel with the STR alleles. The ILS includes DNA fragmentsof known sizes, and can be tagged with a pre-determined fluorescent dye.The detector unit detects fluorescence emitted from the ILS to set up asize scale. In addition, the detector unit detects fluorescence emittedfrom the STR alleles. The detector unit can suitably convert thedetected fluorescence into electrical signals. The electrical signalscan be suitably stored and/or analyzed. In an example, a processorexecutes DNA analysis software instructions to identify the STR allelesby their sizes and emitted fluorescence colors (wavelengths).

The computing module 170 includes computing and communication units. Inan example, the computing module 170 includes a personal computer. Thepersonal computer can be coupled with the controller module 180 toprovide a user interface. The user interface can inform the status ofthe DNA analyzer 100, and can receive user instructions for controllingthe operation of the DNA analyzer 100. The personal computer includesvarious storage media to store software instruction and data. Thepersonal computer can include DNA analysis software that can performdata processing based on raw data obtained from the detection module150. In addition, the personal computer can be coupled to externalprocessing units, such as a database, a server, and the like to furtherprocess the data obtained from the DNA analyzer 100.

The magnetic module 190 can enable a magnetic solid phase for theintegrated single chip DNA analysis. In an embodiment, the magneticsolid phase can be suitably incorporated in the integrated single chipDNA analysis to facilitate a volume reduction to suit for low copynumbers of template DNAs. In another embodiment, the magnetic solidphase can be suitably incorporated into an integrated single chipsequencing DNA analysis.

The controller module 180 can receive status signals and feedbacksignals from the various components, and provide control signals to thevarious components according to a control procedure. In addition, thecontroller module 180 can provide the status signals to, for example,the personal computer, to inform the user. Further, the controllermodule 180 can receive user instructions from the personal computer, andmay provide the control signals to the various components based on theuser instructions.

During operation, the controller module 180 receives user instructionsfrom the personal computer to perform a STR typing analysis, forexample. The controller module 180 then monitors the microfluidic chipmodule 110 to check whether a suitable disposable cartridge has beeninstalled, and whether swabs have been identified and suitably immersedin the liquid phase mixture to extract template DNA. When the controllermodule 180 confirms the proper status at the microfluidic chip module110, the controller module 180 starts a control procedure correspondingto the STR typing analysis. In an example, the controller module 180 cancontrol the thermal module 120 to maintain an appropriate temperature atthe wells of the sample acceptor for a predetermined time. The liquidphase mixture in the wells can extract template DNAs from the swabs.Then, the controller module 180 can control the pressure module 130 topump the extracted template DNAs into the first domain 111 a of themicrofluidic chip 111. In addition, the controller module 180 cancontrol the pressure module 130 to pump reagents for multiplexed STRamplification into the first domain 111 a.

Further, the controller module 180 can control the thermal module 120 toinduce thermal cycling for the multiplexed STR amplification at thefirst domain 111 a. The reagents and the thermal cycling can cause DNAamplification. In addition, the DNA amplicons can be suitably taggedwith fluorescent labels.

Subsequently, the controller module 180 can control the pressure module130 to flow the DNA amplicons to the second domain 111 b. The controllermodule 180 may control the pressure module 130 to pump a dilutionsolution into the microfluidic chip 111 to mix with the DNA amplicons.In addition, the controller module 180 may control the pressure module130 to pump an ILS into the microfluidic chip 111 to mix with the DNAamplicons.

Further, the controller module 180 controls the high voltage module 140to induce electro-kinetic injection to inject DNA fragments into themicro channels. The DNA fragments include the amplified targets, and theILS. Then, the controller module 180 controls the high voltage module140 to induce electrophoretic separation in the micro channels.Additionally, the controller module 180 can control the thermal module120 to maintain a suitable temperature at the second domain 111 b duringseparation, for example, to maintain the temperature for denaturingseparation of the DNA fragments.

The controller module 180 then controls the detection module 150 todetect the labeled DNA fragments. The detection module 150 can emit anddirect a laser beam to the micro channels to excite the fluorescentlabels to emit fluorescence. Further, the detection module 150 candetect the emitted fluorescence and store detection data in a memory.The detection data can include a detection time, and a detected color(wavelength), along with a detected intensity, such as a relativemagnitude of the detected fluorescence. The detection data can betransmitted to the personal computer for storage. Additionally, thecontroller module 180 can provide control statuses to the personalcomputer to inform the user. For example, the controller module 180 cansend an analysis completed status to the personal computer when thecontrol procedure is completed.

The DNA analyzer 100 can be suitably configured for various DNA analysesby suitably adjusting the reagents housed by the reagent carrier and thecontrol procedure executed by the controller module 180.

FIG. 2A shows a swab storage example 212, and FIGS. 2B-2C show a sideelevation view and a front elevation view of a sample cartridge example215 according to an embodiment of the disclosure. The swab storage 212includes a case 203, a seal cap 202 and a swab 205. The seal cap 202 andthe swab 205 are attached together. In addition, the swab storage 212includes an identifier, such as a barcode label 204 that can be attachedto the case 203, an RFID tag 201 that can be implanted in the seal cap202, and the like.

Before taking DNA sample, the swab 205 is safely stored in the case 203to avoid contamination. After taking DNA sample, the swab 205 can beplaced in the sample cartridge 215.

The sample cartridge 215 can include a microfluidic chip 211, a sampleacceptor 207 and a reagent carrier 206. The sample acceptor 207 includesa plurality of separated wells 207A-207D for taking swabs. Each wellincludes a liquid phase mixture 214 that is sealed by a membrane 208 ata bottom portion of the well. The liquid phase mixture 214 can conductenzymatic digestion of all proteins and other cellular interferences,with the exception of DNA, and thus can perform DNA extraction andpurification when a swab with DNA sample is inserted in the liquid phasemixture 214.

While the sample cartridge 215 is described in the context of swabs, itshould be understood that the sample cartridge 215 can be suitablyadjusted to suit other DNA gathering methods, such as blood stain cards,airborne samples, fingerprints samples, and the like.

In an embodiment, the seal cap 202 is a stepped seal cap that can sealthe well in a first step, and a second step. When the seal cap 202 sealsthe well in the first step, the swab 205 does not puncture the membrane208, and can be safely sealed in the well to maintain sample integrity.When the seal cap 202 seals the well in the second step, the swab 205punctures the membrane 208 and is immersed in the liquid phase mixture214.

The reagent carrier 206 houses various solutions for DNA analysis. In anSTR typing example, the reagent carrier houses reagents for multiplexedSTR amplification. In addition, the reagent carrier houses a coatingsolution, such as poly-N-hydroxyethylacrylamide, and the like. Thecoating solution can be used to coat micro channel walls prior to theseparation. Further, the reagent carrier houses a dilution solution,such as water, formamide, and the like. The dilution solution can beused to reduce the ionic strength in order to promote betterelectro-kinetic injection. In an embodiment, the reagent carrier housesan internal lane standard (ILS). The ILS can be used for sizemeasurement. The reagent carrier also houses a polymer solution forelectrophoretic separation in the micro-scale chip environment.

During operation, for example, a new disposable cartridge 215 is takenfrom a storage package, and installed in a DNA analyzer, such as the DNAanalyzer 100. Then, a swab 205 can be used to take a DNA sample. Theswab 205 is then identified and inserted into one of the wells 207A-207Dand sealed in the first step. Additional swabs 205 can be used to takeDNA samples, and then identified and inserted into the un-used wells207A-207D. Further, the DNA analyzer 100 can include a mechanism thatcan push the seal caps 202 to seal the wells 207A-207D in the secondstep, thus the swabs 205 can puncture the membrane 208, and immerse inthe liquid phase mixture 214.

FIG. 3 shows a schematic diagram of a microfluidic chip example 311according to an embodiment of the disclosure. The microfluidic chip 311includes various micro structures, such as inlets 312-314, reactionreservoirs 315-316, channels 317 a-317 b, electrode reservoirs 318,outlets (not shown), and the like, that are integrated for single-chipDNA analysis. It is noted that the various micro structures can bedesigned and integrated to suit for various DNA analyses, such as STRtyping, sequencing, and the like.

The inlets 312-314 can be coupled to a pressure module to injectsolutions in the microfluidic chip 311. As described above, theconnection can be made via a micro-macro interface. In an example, theinlet 312 is for injecting a template DNA solution from a well of thesample acceptor 207, and the inlet 313 is for injecting PCR reagentsfrom the reagent carrier 206. In addition, the inlet 313 can be used forinjecting dilution solution and ILS from the reagent carrier 206.

The reaction reservoirs 315-316 are configured for various purposes. Inan example, the reaction reservoir 315 is configured for the PCRamplification, and the reaction reservoir 316 is configured for thepost-PCR processes, such as dilution, and the like. More specifically,the reaction reservoir 315 is located in a first domain 311 a, which isa thermal control domain. The temperature within the thermal controldomain 311 a can be precisely controlled. In an example, an infraredheating unit directs heat to the thermal control domain 311 a, a coolingfan disperses heat from the thermal control domain 311 a, and aninfrared sensing unit measures a temperature in the thermal controldomain 311 a. The infrared heating unit and the cooling fan can becontrolled based on the temperature measured by the infrared sensingunit. The infrared heating unit, the cooling fan, and the infraredsensing unit can perform thermal control without contacting the thermalcontrol domain 311 a.

In another example, the temperature in the thermal control domain 311 ais measured by a thermal coupling technique. More specifically, themicrofluidic chip 311 includes a thermal-coupler reservoir 319 withinthe first domain 311 a. Thus, the solution temperature within thereaction reservoir 315 and the thermal-coupler reservoir 319 can beclosely related.

The solution temperature within the thermal-coupler reservoir 319 can bemeasured by any suitable technique. Based on the measured solutiontemperature within the thermal-coupler reservoir 319, the solutiontemperature within the reaction reservoir 315 can be determined. Then,the infrared heating unit and the cooling fan can be controlled based onthe temperature measured by the thermal coupling technique in order tocontrol the solution temperature in the reaction reservoir 315.

In-an embodiment, after the PCR amplification, the PCR mixture isfluidically directed from the reaction reservoir 315 to a post-PCRclean-up/dilution domain, such as the reaction reservoir 316. In thereaction reservoir 316, the PCR mixture is diluted. In an example, thePCR mixture and a dilutant solution are mixed together according to aratio from 1:5 to 1:20 (1 part of PCR mixture to 5-20 parts ofdilutant). Further, ILS can be added in the reaction reservoir 316 tomix with the PCR mixture.

The channels 317 a-317 b are located in a second domain 311 b. Electricfields can be suitably applied onto the channels 317 a-317 b. In anexample, the channels 317 a-317 b are configured according to a cross-Tdesign, having a short channel 317 a and a long channel 317 b.

The electrode reservoirs 318 can be used to apply suitable electricfields over the short channel 317 a and the long channel 317 b. Thus,the short channel 317 a is configured for electro-kinetic injection, andthe long channel 317 b is configured for electrophoretic separation. Forexample, when a high voltage is applied to the short channel 317 a, DNAfragments can be injected from the reaction reservoir 316 into the shortchannel 317 a at the intersection of the short channel 317 a and thelong channel 317 b. The long channel 317 b can be filed with sievingmatrix. When a high voltage is applied to the long channel 317 b, theinjected DNA fragments can migrate in the long channel 317 b to thepositive side of the electric field induced by the high voltage, in thepresence of the sieving matrix. In an example, the length of the longchannel 317 b is about 8.8 cm with detection at about 8 cm from theintersection.

It should be understood that the microfluidic chip 311 can include otherstructures to assist DNA analysis. In an example, the microfluidic chip311 includes an alignment mark 321. The alignment mark 321 can assist adetection module to align to the long channel 317 b.

During operation, for example, the inlet 312 can input a template DNAinto the reaction reservoir 315, and the inlet 313 can input PCRreagents into the reaction reservoir 315. Then, thermal-cycling can beinduced at the first domain 311 a, and PCR amplification can beconducted in the reaction reservoir 315 to amplify DNA fragments basedon the template DNA and the PCR reagents. After the PCR amplification,the DNA amplicons in the reaction reservoir 315 can be mobilized intothe reaction reservoir 316 in a liquid flow. In the reaction reservoir316, a dilution solution and ILS can be input to mix with the DNAfragments. Further, the DNA fragments in the reaction reservoir 316 canbe injected across the short channel 317 a by electro-kinetic injection.The DNA fragments then migrate in the long channel 317 b under the forceof electric field applied over the long channel 317 b. The speed ofmigration depends on the sizes of the DNA amplicons, in the presence ofthe sieving matrix. Thus, the DNA fragments are separated in the longchannel 317 b according to their sizes.

FIG. 4 shows an exemplary DNA analyzer 400 according to an embodiment ofthe disclosure. The DNA analyzer 400 is packaged in a box. The boxincludes handles, wheels and the like, to facilitate transportation ofthe DNA analyzer 400. In an implementation, the total weight of the DNAanalyzer 400 is less than 70 lb, and is appropriate for two persons tocarry.

The DNA analyzer 400 is implemented in a modular manner. Each module canbe individually packaged, and can include an interface for inter-modulecouplings. Thus, each module can be easily removed and replaced. Themodular design can facilitate assembly, troubleshooting, repair, and thelike.

The DNA analyzer 400 includes a user module (UM) 410, an active pressuremodule (APM) 430, a detection module 450, a power module (PM) 460, acomputing module 470, and a controller module (CM) 480. In addition, theDNA analyzer 400 includes a sample cartridge storage 415 and a swabstorage 412.

The UM 410 includes a holder to hold a sample cartridge, such as thesample cartridge 215, at an appropriate position when the samplecartridge is inserted by a user. Further, the UM 410 includes interfacecomponents to couple the sample cartridge 215 with, for example, the APM430, the detection module 450, and the like. The UM 410 includes thermalcomponents, such as resistance heaters 421, a cooling fan 422, aninfrared heating unit 423, and the like. The thermal components can besuitably positioned corresponding to the sample cartridge 215. Forexample, a resistance heater 421 is situated at a position that caneffectively control a temperature of the liquid phase mixture within theplurality of separated wells on the sample cartridge 215. Thetemperature can be determined to optimize enzyme activities of theliquid phase mixture to conduct enzymatic digestion of all proteins andother cellular interferences, with the exception of DNA. Anotherresistance heater 421 is at a position that can effectively control atemperature of the separation channel on the microfluidic chip 211. Theinfrared heating unit is at a position that can direct heat to thethermal control domain of the microfluidic chip 211 on the samplecartridge 215. The cooling fan is at a position that can effectivelydisperse heat from the thermal control domain. Further, the UM 410includes a high voltage module that can apply suitable high voltages viathe electrode reservoirs of the microfluidic chip 211.

It is noted that the UM 410 can include other suitable components. In anembodiment, the UM 410 includes a magnetic module that can suitablyapply magnetic control over a domain of the microfluidic chip 211.

The APM 430 includes suitably components, such as pumps, vacuums, andthe like, to apply suitable pressures to the microfluidic chip 211 toenable fluid movement.

The PM 460 receives an input main power, and generates various operationpowers, such as 6 V, 12 V, 24 V, 1000V, 2000V, and the like, for variouscomponents of the DNA analyzer 400.

The detection module 450 can include a laser module (LM) 451, a passiveoptics module (POM) 452, and an active optics module (AOM) 453. The LM451 can include any suitable device to emit a laser beam. In anembodiment, the LM 451 includes an argon-ion laser. In another example,the LM 451 includes a diode laser. In another embodiment, the LM 451includes a solid state laser, such as a coherent sapphire opticallypumped semiconductor laser. The solid state laser can have a reducedsize and weight, and can consume less power than the argon-ion laser. Inaddition, the solid state laser generates less waste heat, such that fansize can be reduced to reduce footprint of the DNA analyzer 400.

The AOM 453 includes optical elements that may need to be adjusted withregard to each inserted microfluidic chip. In an example, the AOM 453includes a plurality of optical fibers that are respectively coupled toa plurality of separation channels on the microfluidic chip. Theplurality of optical fibers can respectively provide laser beams to theplurality of separation channels to excite fluorescence emission. Inaddition, the plurality of optical fibers can return the emittedfluorescence from the plurality of separation channels.

The POM 452 includes various optical elements, such as lens, splitters,photo-detectors, and the like, that do not need to be adjusted withregard to each inserted microfluidic chip. In an example, the POM 452 iscalibrated and adjusted with regard to the LM 451 and the AOM 453 whenthe detection module 450 is assembled. Then, the optical elements withinthe POM 452 are situated at relatively fixed positions, and do not needto be adjusted with regard to each inserted microfluidic chip.

The controller module 480 is coupled to the various components of theDNA analyzer 400 to provide control signals for DNA analysis. Thecontroller module 480 includes a control procedure that determinessequences and timings of the control signals.

The computing module 470 is implemented as a personal computer. Thepersonal computer includes a processor, a memory storing suitablesoftware, a keyboard, a display, and a communication interface. Thecomputing module 470 can provide a user interface to ease user controland monitor of the DNA analysis by the DNA analyzer 400.

FIG. 5 shows a flow chart outlining a process example for using a DNAanalyzer, such as the DNA analyzer 400, to perform DNA analysisaccording to an embodiment of the disclosure. The process starts atS501, and proceeds to S510.

At S510, a user of the DNA analyzer 400 plugs in a main power supply. Inan embodiment, the main power supply can be a 110 V, 50 Hz, AC powersupply, or can be a 220V, 60 Hz, AC power supply. The power module 460can convert the main power supply to a plurality of operation powers,and provide the plurality of operation powers to the various modules ofthe DNA analyzer 400. Then, the process proceeds to S515.

At S515, the user starts up a user control interface. For example, theuser turns on the personal computer 470, and starts a software packagethat interacts with the user and the controller module 480. The softwarepackage enables the personal computer 470 to provide a user controlinterface on the display. Further, the software package enables thepersonal computer 470 to receive user instructions via the keyboard ormouse. The software packages can also enable the personal computer 470to communicate with the controller module 480. Then, the processproceeds to S520.

At S520, the user instructs the DNA analyzer 400 to initialize. The usercontrol interface receives the initialization instruction, and thesoftware package enables the personal computer 470 to send theinitialization instruction to the controller module 480. The controllermodule 480 can then initialize the various components of the DNAanalyzer 400. For example, the controller module 480 can power on thevarious components, check the status and reset the status if needed.Then, the process proceeds to S525.

At S525, the user inserts a sample cartridge 215 in the UM 410. Thesample cartridge 215 can be positioned by a holder. The interfacecomponents can suitably couple the sample cartridge 215 to othercomponents of the DNA analyzer 400. Then, the process proceeds to S530.

At S530, the user takes a swab 205, and lets the DNA analyzer 400 toidentify the swab 205. In an example, the DNA analyzer 400 includes abarcode reader that can read the barcode label 204 attached to the case203 for storing the swab 205. In another example, the DNA analyzer 400excites the RFID 201 implanted in the seal cap 202 of the swab 205 toobtain a unique serial number of the swab 205. Then, the processproceeds to S535.

At S535, the user uses the swab 205 to take a DNA sample and inserts theswab 205 into a well of the sample cartridge 215. The user may repeatthe steps S530 and S535 to insert multiple swabs 205 into the separatedwells of the sample cartridge 215. Then, the process proceeds to S540.

At S540, the user instructs the DNA analyzer 400 to start a DNAanalysis. The user control interface receives the start instruction, andthe software package enables the personal computer 470 to send the startinstruction to the controller module 480. The controller module 480 canstart a control procedure corresponding to the DNA analysis. In anexample, the controller module 480 starts an STR typing procedurecorresponding to a multiplexed STR typing analysis. In another example,the controller module 480 starts a sequencing procedure corresponding toDNA sequencing analysis. Then, the process proceeds to S545.

At S545, the user waits and monitors the status of the DNA analysis. Thecontrol procedure can specify sequences and timings of control signalsto various components of the DNA analyzer 400 corresponding to the DNAanalysis. Then, the controller module 480 automatically sends thecontrol signals according to the sequences and the timings specified inthe control procedure. In addition, the controller module 480 receivesstatus and feedback signals from the various components, and sends themto the personal computer 470. The personal computer 470 then providesthe analysis status for the user to monitor. Then, the process proceedsto S550.

At S550, the controller module 480 finishes executing the controlprocedure, and sends an analysis-completed status to the personalcomputer 470. The personal computer 470 can inform the user of theanalysis-completed status via the user control interface. Then, theprocess proceeds to S555.

At S555, the user performs post data processing. The user can store theraw data of the DNA analysis, or transmit the raw data to a remotereceiver. In addition, the user may start a software package for postdata processing. Alternatively, the software package for post dataprocessing can be suitably integrated with the control procedure. Thus,after the control procedure is successfully executed, the softwarepackage for post data processing is executed automatically to performpost data processing. The process then proceeds to S599 and terminates.

It is noted that to perform another DNA analysis, the user may throwaway the sample cartridge and repeat S520-S550. It is also noted thatthe sequence of the DNA analysis steps can be suitably adjusted. Forexample, S535 and S530 can be swapped, thus a swab can be first used totake a DNA sample, and then identified by the DNA analyzer 400.

FIG. 6 shows a flow chart outlining a process example 600 for a DNAanalyzer to perform multiplexed STR typing according to an embodiment ofthe disclosure. The process starts at S601 and proceeds to S610.

At S610, the controller module 480 controls the resistance heater 421 tomaintain a temperature for template DNA extraction and purification.More specifically, the resistance heater 421 is positioned correspondingto the plurality of wells on the sample cartridge 215. A well can accepta swab 205. The swab 205 can puncture the membrane that seals the liquidphase mixture at the bottom of the well, thus the swab 205 is immersedinto the liquid phase mixture. The liquid phase mixture can extract andpurify a template DNA from the swab at the temperature according toenzymatic DNA isolation method. In an embodiment, the liquid phasemixture can achieve a compatible DNA concentration and purity to silicabased solid phase extraction method in about 6 minutes. Then, theprocess proceeds to S620.

At S620, the controller module 480 controls the APM 430 to flow theextracted template DNA and reagents to a reaction reservoir for the PCRamplification. For example, the reagent carrier 206 houses reagents formultiplexed STR amplification. The controller module 480 sends controlsignals to the APM 430. In response to the control signals, a pump pumpsthe liquid phase mixture from the well to the reaction reservoir, andanother pump pumps the reagents from the reagent carrier 206 to thereaction reservoir. Then, the process proceeds to S630.

At S630, the controller module 480 controls the cooling fan 422 and theinfrared heating unit 423 to induce thermal cycling in the reactionreservoir for the multiplexed STR amplification. In addition, thereagents can attach fluorescent labels to the DNA amplicons during theSTR amplification process. The process then proceeds to S640.

At S640, after the PCR amplification, the solution can be diluted. Morespecifically, the controller module 480 sends control signals to the APM430 after the PCR amplification. In response to the control signals, theAPM 430 flows the DNA amplicons into a dilution reservoir. In addition,the APM 430 flows a dilution solution from the reagent carrier into thedilution reservoir. The process then proceeds to 5650.

At S650, the controller module 480 sends control signals to the highvoltage module in the UM 410 to inject the DNA amplicons across theinjection arm (the short channel 317 a). Then, the process proceeds toS660.

At S660, the controller module 480 sends control signals to the highvoltage module in the UM 410 to apply appropriate high voltage over theseparation channel (the long channel 317 b) to separate the DNAamplicons based on sizes. The process then proceeds to S670.

At S670, the controller module 480 sends control signals to thedetection module 450 to excite the fluorescent labels to emitfluorescence and detect the emitted fluorescence. The raw detection datacan be sent to the personal computer 470 for storage andpost-processing. The process then proceeds to S699, and terminates.

It is noted that some process steps in the process 600 can be executedin parallel. For example, the step S660 and the step S670 can beexecuted in parallel. The controller module 480 sends control signals toboth the high voltage module in the UM 410 and the detection module 450at about the same time. The control signals to the high voltage modulein the UM 410 cause the electrophoretic separation in the separationchannel, while the control signals to the detection module 450 causefluorescence detection.

It is noted that the process 600 can be suitably adjusted along withreagents adjustments for other DNA analysis, such as qPCR DNAquantitation, sequencing, and the like.

In a qPCR DNA quantitation example, step S601 to S630 are executed, andstep S640 to S670 can be deleted. In addition, in step S630, whenthermal cycles are induced in a qPCR reservoir for PCR amplification,the controller module 480 sends control signals to the detection module450 to detect florescence emitted by the fluorescent labels in the qPCRreservoir.

It is also noted that a magnetic solid phase purification process stepcan be suitably added into the process 600 to facilitate further volumereduction, thus the process 600 can be adjusted for DNA sequencing.

FIG. 7 shows a plot of thermal cycling for polymerase chain reaction(PCR) amplification induced in two systems. Thermal cycling 710 isinduced in a first system, and thermal cycling 720 is induced in asecond system. Each of the thermal cycling 710 and the thermal cycling720 is induced by an infrared light source and a cooling fan based on atemperature measurement using a thermal-coupler reservoir technique.Both systems achieve a temperature fluctuation of less than ±0.1° C. Inaddition, a system-to-system reproducibility is achieved.

FIG. 8 shows a plot 800 of a 16-loci STR typing analysis result obtainedfrom an implemented DNA analyzer according to an embodiment of thedisclosure. The DNA analyzer accepts a sample, such as in the form of aswab, extracts a template DNA from the sample, and acts on amicrofluidic chip to perform an integrated single-chip DNA analysisbased on the extracted template DNA. More specifically, the DNA analyzerpumps the extracted template DNA and 16-loci STR reagents in a reservoirwithin a first domain of the microfluidic chip. Then, the DNA analyzerinduces thermal cycling in the first domain of the microfluidic chip toperform multiplexed 16-loci PCR amplification based on the template DNA.The 16-loci STR reagents use four fluorescent labels having differentwavelengths to label the DNA fragments. After the PCR amplification, theDNA analyzer pumps ILS into the amplified STR alleles. The ILS islabeled with a fifth fluorescent label having a different wavelengthfrom the four fluorescent labels used by the 16-loci STR reagents.Further, the DNA analyzer injects the STR alleles along with the ILSinto a second domain of the microfluidic chip. The second domainincludes a separation channel. The DNA analyzer induces an electricalfield over the separation channel for electrophoretic separation. Underthe electrical field, the STR alleles and the ILS migrate in theseparation channel based on sizes. The DNA analyzer then generates anddirects a laser beam to the separation channel to excite the fluorescentlabels to emit fluorescence. Further, the emitted fluorescence iscollected and detected by the DNA analyzer. The DNA analyzer detectsfluorescence intensities of different wavelengths over time, andidentifies an STR in the sample based on a combination of wavelength andsize comparison with ILS.

The plot 800 shows five curves 810-850. The curve 810 shows detectedfluorescence intensity versus time for a first wavelength. The curve 820shows detected fluorescence intensity versus time for a secondwavelength. The curve 830 shows detected fluorescence intensity versustime for a third wavelength. The curve 840 shows detected fluorescenceintensity versus time for a fourth wavelength, and the curve 850 showsdetected fluorescence intensity versus time for a fifth wavelength. Thefifth wavelength corresponds to the fluorescent label used to tag theILS, thus spikes in the curve 850 are of known sizes. Spikes in thecurves 810-840 correspond to STR alleles in the sample under analysis.The sizes of the STR alleles can be determined based on comparison tothe spikes in the curve 850. Thus, the STR alleles can be identifiedbased on the wavelength and the size comparison with the ILS.

While the invention has been described in conjunction with the specificexemplary embodiments thereof, it is evident that many alternatives,modifications, and variations will be apparent to those skilled in theart. Accordingly, exemplary embodiments of the invention as set forthherein are intended to be illustrative, not limiting. There are changesthat may be made without departing from the spirit and scope of theinvention.

1. A microfluidic chip, comprising: a first domain configured forpolymerase chain reaction (PCR) amplification of DNA fragments; adilution domain that is fluidically coupled to the first domain and isconfigured to dilute a PCR mixture received from the first domain with adilutant; and a second domain that is fluidically coupled to thedilution domain so as to receive the amplified DNA fragments, the seconddomain including a separation channel that is configured to performelectrophoretic separation of the amplified DNA fragments.
 2. Themicrofluidic chip of claim 1, wherein the first domain comprises: areservoir configured to receive a template DNA and reagents, and amplifythe DNA fragments based on the template DNA and the reagents.
 3. Themicrofluidic chip of claim 2, further comprising: a plurality of inletsconfigured to input the template DNA and the reagents into themicrofluidic chip.
 4. The microfluidic chip of claim 1, wherein thesecond domain comprises: an injection channel configured to inject theamplified DNA fragments into the separation channel by electro-kineticinjection.
 5. The microfluidic chip of claim 1, further comprising: aplurality of electrode reservoirs for applying an electric field overthe separation channel.
 6. The microfluidic chip of claim 1, wherein inthe dilution domain, the PCR mixture and the dilutant are mixedaccording to a ratio from 1:5 to 1:20.
 7. A DNA analyzer that isconfigured to receive and act on the microfluidic chip of claim 1 toperform DNA analysis.
 8. A cartridge, comprising: a sample acceptorconfigured to extract a template DNA; a microfluidic chip having a firstdomain and a second domain coupled to the first domain, wherein thefirst domain is configured to perform polymerase chain reaction (PCR)amplification of DNA fragments based on the template DNA, and the seconddomain has a separation channel that is configured to performelectrophoretic separation of the amplified DNA fragments.
 9. Thecartridge of claim 8, further comprising: a reagent carrier configuredto carry reagents for the PCR amplification, and solutions for theelectrophoretic separation.
 10. The cartridge of claim 8, wherein thesample acceptor is configured to extract the template DNA by at leastone of a silica solid phase extraction, and a liquid phase enzymatic DNAisolation.
 11. The cartridge of claim 8, wherein the sample acceptorcomprises: a well having a liquid phase mixture to extract the templateDNA.
 12. The cartridge of claim 11, wherein the liquid phase mixture issealed in the well before the extraction.
 13. The cartridge of claim 9,wherein the first domain of the microfluidic chip further comprises: areservoir configured to receive the extracted template DNA and thereagents, and amplify the DNA fragments based on the template DNA andthe reagents.
 14. The cartridge of claim 9, wherein the microfluidicchip further comprises: a plurality of inlets configured to input thetemplate DNA and the reagents kits into the microfluidic chip.
 15. Thecartridge of claim 8, wherein the second domain of the microfluidic chipcomprises: an injection channel configured to inject the amplified DNAfragments into the separation channel by electro-kinetic injection. 16.The cartridge of claim 8, wherein the microfluidic chip furthercomprises: a plurality of electrode reservoirs for applying an electricfield over the separation channel.
 17. The cartridge of claim 8, whereinthe microfluidic chip further comprises: a waste outlet to drain liquidout of the microfluidic chip.
 18. The cartridge of claim 8, wherein themicrofluidic chip further comprises: a dilution domain coupled to thefirst domain and the second domain, the dilution domain diluting a PCRmixture received from the first domain with a dilutant, and providingthe diluted PCR mixture to the second domain.
 19. The cartridge of claim18, wherein in the dilution domain, the PCR mixture and the dilutant aremixed according to a ratio from 1:5 to 1:20.
 20. A DNA analyzer that isconfigured to receive the cartridge of claim 8, and act on themicrofluidic chip to perform DNA analysis.
 21. A DNA analyzer,comprising: an interface for coupling a microfluidic chip to the DNAanalyzer, wherein the microfluidic chip includes: a first domainconfigured for polymerase chain reaction (PCR) amplification of DNAfragments, the DNA fragments being labeled with fluorescent labels; anda second domain that is coupled to the first domain so as to receive theamplified DNA fragments, the second domain including a separationchannel that is configured to perform electrophoretic separation of theamplified DNA fragments; a pressure module configured to flow liquid inthe microfluidic chip; a thermal module configured to induce thermalcycles at the first domain of the microfluidic chip for the PCRamplification; a power module configured to generate voltages to beapplied to the second domain of the microfluidic chip for theelectrophoretic separation; a detection module configured to excite thefluorescent labels to emit fluorescence, and detect the emittedfluorescence; and a controller module configured to control the pressuremodule, the thermal module, the power module, and the detection moduleaccording to a control procedure to act on the microfluidic chip for asingle-chip DNA analysis.
 22. The DNA analyzer of claim 21, wherein thepressure module further comprises: a plurality of pumps configured toinject a template DNA and reagents in the microfluidic chip.
 23. The DNAanalyzer of claim 22, wherein the controller module is configured torespectively control the plurality of pumps.
 24. The DNA analyzer ofclaim 21, wherein the pressure module further comprises: a vacuum pumpconfigured to control a membrane valve on the microfluidic chip.
 25. TheDNA analyzer of claim 21, wherein the thermal module further comprises:a heating unit configured to direct heat to the first domain; a coolingunit configured to disperse heat from the first domain; and a sensingunit configured to measure a temperature at the first domain.
 26. TheDNA analyzer of claim 25, wherein the heating unit includes an infraredlight source to heat the first domain.
 27. The DNA analyzer of claim 25,wherein the sensing unit includes an infrared pyrometer to measure thetemperature.
 28. The DNA analyzer of claim 25, wherein the sensing unitmeasures a temperature within a thermal coupler reservoir in the firstdomain, the thermal coupler reservoir being thermally coupled with areaction reservoir in the first domain configured for the PCRamplification.
 29. The DNA analyzer of claim 21, wherein the detectionmodule further comprises: a laser module configured to emit a laserbeam; a detection module configured to detect fluorescence; and a set ofoptics to direct the laser beam to the separation channel to excite thefluorescent labels to emit fluorescence, and direct the excitedfluorescence to the detection module for detection.
 30. The DNA analyzerof claim 21, further comprising at least one of: a barcode readerconfigured to read a barcode to identify a DNA sample; and a radiofrequency identification (RFID) reader configured to read a RFID tag toidentify a DNA sample.
 31. The DNA analyzer of claim 21, furthercomprising: a magnetic module configured to control magnetic beads usedin a magnetic solid phase of a DNA analysis process.
 32. The DNAanalyzer of claim 21, wherein the microfluidic chip further comprises: adilution domain coupled to the first domain and the second domain, thedilution domain diluting a PCR mixture received from the first domainwith a dilutant, and providing the diluted PCR mixture to the seconddomain.
 33. The DNA analyzer of claim 32, wherein in the dilutiondomain, the PCR mixture and the dilutant are mixed according to a ratiofrom 1:5 to 1:20.
 34. A method for DNA analysis, comprising: inducingthermal cycles in a first domain of a microfluidic chip for PCRamplification of DNA fragments; inducing liquid flow to move theamplified DNA fragments from the first domain to a second domain of themicrofluidic chip having a separation channel for electrophoreticseparation; inducing an electric field over the separation channel toseparate the DNA fragments by sizes; and detecting the separated DNAfragments.
 35. The method of claim 34, further comprising: maintaining atemperature for enzymatic DNA extraction to extract a template DNA. 36.The method of claim 35, further comprising: injecting reagents into areservoir in the first domain for the PCR amplification; and injectingthe template DNA into the reservoir for the PCR amplification.
 37. Themethod of claim 34, further comprising: maintaining a temperature in theseparation channel.
 38. The method of claim 34, wherein inducing theliquid flow to move the amplified DNA fragments from the first domain tothe second domain of the microfluidic chip having the separation channelfor electrophoretic separation, further comprises: diluting theamplified DNA fragments with a dilution solution; and electro-kineticinjecting the DNA fragments into the separation channel.
 39. The methodof claim 34, wherein the amplified DNA fragments are tagged withfluorescent labels, and detecting the separated DNA fragments furthercomprises: emitting a laser beam; directing the laser beam to theseparation channel to excite the fluorescent labels to emitfluorescence; collecting the emitted fluorescence; and returning thecollected fluorescence for detection.
 40. The method of claim 38,further comprising: adding an internal lane standard (ILS) with theamplified DNA fragments to assist a size measurement.
 41. The method ofclaim 38, wherein diluting the amplified DNA fragments with the dilutionsolution further comprises: diluting a PCR mixture received from thefirst domain with the dilution solution according to a ratio of 1:5 to1:20 (one part of PCR mixture to 5-20 parts of dilution solution).